Electrochemical-sensor design

ABSTRACT

An electrochemical test sensor adapted to assist in determining the concentration of analyte in a fluid sample is disclosed. The sensor comprises a base that assists in forming an opening for introducing the fluid sample, a working electrode being coupled to the base, and a counter electrode being coupled to the base, the counter electrode and the working electrode being adapted to be in electrical communication with a detector of electrical current, and a sub-element being coupled to the base. A major portion of the counter electrode is located downstream relative to the opening and at least a portion of the working electrode. The sub-element is located upstream relative to the working electrode such that when electrical communication occurs between only the sub-element and the working electrode there is insufficient flow of electrical current through the detector to determine the concentration of the analyte in the fluid sample.

This is a Divisional application of co-pending application Ser. No.09/861,437 filed May 21, 2001, now U.S. Pat. No. 6,841,052 which in turnis a Continuation-In-Part of application Ser. No. 09/731,943 filed Dec.8, 2000, now U.S. Pat. No. 6,531,040 B2, which is in turn aContinuation-In-Part of application Ser. No. 09/366,269, filed on Aug.2, 1999, now abandoned.

BACKGROUND OF THE INVENTION

The present invention relates to an electrochemical biosensor that canbe used for the quantitation of a specific component (analyte) in aliquid sample. Electrochemical biosensors of the type underconsideration are disclosed in U.S. Pat. Nos. 5,120,420 and 5,264,103.These devices have an insulating base upon which carbon electrodes areprinted with the electrodes being covered with a reagent layercomprising a hydrophilic polymer in combination with an oxidoreductasespecific for the analyte. These patents typically involve a spacerelement, a generally U shaped piece and a cover piece, so that when thebase, spacer element and cover piece are laminated together, there iscreated a capillary space containing the electrodes covered by thereagent layer. In addition to the oxidoreductase, there is included anelectron acceptor on the reagent layer or in another layer within thecapillary space. A hydrophilic polymer, e.g. carboxymethylcellulose, isused to facilitate the drawing of the aqueous test fluid into thecapillary space.

In U.S. Pat. No. 5,141,868 there is disclosed another sensor in whichthe electrodes are contained within a capillary space. This referencedescribes the method of preparing a sensor by mating the base and coverplates which are adhered to the base to form a capillary space intowhich a fluid test sample such as blood is drawn. An alternative to thisdesign is disclosed in U.S. Pat. No. 5,798,031 in which the sensor iscomprised of two pieces, a base and a concave lid which, when fusedtogether, form the capillary space. In either embodiment, working andcounter electrodes are screen printed onto the base so that anelectrochemically created current can flow when these electrodes areelectrically connected and a potential created between them.

These devices have a base plate and lid which are laminated togetherwith the U shaped spacer element in between so that the U shaped portionis open to provide a capillary space between the base and the cover.Touching the opening in the side of the sensor to a drop of test fluidsuch as blood results in the blood being drawn into the capillary space,so that it covers the reaction layer on the surface of the workingelectrode. An enzymatic reaction between the oxidoreductase creates aflow of electrons which are carried by a mediator such as ferricyanideto the working electrode and flow through the working electrode to ameter which measures the magnitude of the current flow. The counterelectrode serves several purposes. First, it provides a fixed potentialagainst which the working electrode is controlled. Second, for a twoelectrode system, such as that depicted in FIGS. 1 and 2, the counterelectrode is used to complete the electrical circuit. In this mode, eachelectron that is transferred to the working electrode is returned to thetest solution on the counter electrode side. The device's software isprogrammed to correlate the magnitude of this flow with theconcentration of analyte in the test sample. In order for this currentto flow, a complete circuit is formed by covering both electrodes withthe conductive test fluid and applying a potential therebetween.

A problem which is sometimes associated with this sort of sensor occurswhen an insufficient amount of blood is applied to the opening so thatthe counter and working electrodes are not completely covered with thesample, resulting in an incomplete current flowing across theelectrodes. Since the amount of analyte such as glucose detected by thesensor is directly portional to the current flowing through thedetection meter, failure to completely cover the sensor's electrodes canresult in an artificially low reading of the blood sample's analyte,e.g., glucose concentration. One technique for dealing with this underfilling problem is disclosed in U.S. Pat. No. 5,628,890 which involves amechanism for preventing any response from being detected when thesample volume is too low to provide an accurate reading. This deviceinvolves a strip comprising an elongated electrode support defining asample transfer path for directional flow of the sample from a sampleapplication point. There is placed a working electrode in the sampletransfer path and a counter or reference electrode down stream from theworking electrode in the sample transfer path. Failure of the bloodsample to totally cover the working electrode will result in no responsefrom the reading mechanism due to the absence of a closed circuitthrough which current can flow. Another technique for detecting shortfills is disclosed in U.S. Pat. No. 5,582,697 where there is described athird electrode located downstream from the working and counterelectrode, so that the circuit between the three electrodes will not becompleted in the event of a short fill.

It would be desirable and it is an object of the present invention toprovide an electrochemical sensor which affirmatively notifies the userwhen insufficient sample has contacted the electrodes. Upon receivingsuch a notice the user knows that an accurate reading cannot be obtainedand that the sensor should be discarded in favor of a new one.

SUMMARY OF THE INVENTION

The present invention is an electrochemical sensor for detecting theconcentration of an analyte, e.g. glucose, in a fluid test sample, suchas blood. The sensor comprises:

-   -   1) a base which provides a flow path for the fluid test sample        having on its surface a counter electrode and a working        electrode in electrical communication with a detector of        electrical current,    -   2) a reaction layer on the surface of at least the working        electrode which contains an enzyme which reacts with the analyte        to produce electrons that are transferred to the working        electrode, and    -   3) a cover which when mated with the base member forms a        capillary space with an opening for the introduction of fluid        test sample into this space. The capillary space encloses the        flow path for the fluid test sample in which the counter and        working electrodes are contained. These electrodes are situated        on the base in relation to the opening so that a major portion        of the counter electrode is located downstream of the opening        from the working electrode. The counter electrode contains a        sub-element which is located upstream of the working electrode,        so that when electrical communication between only the        sub-element of the counter electrode and working electrode due        to incomplete filling of the capillary space by the fluid test        sample occurs, there is insufficient flow of electrical current        through the detector to constitute a valid test for the        concentration of analyte in the fluid test sample. In the event        of such insufficient flow of electrical current, the detector        gives an error signal to notify the user that the test has        failed and that it should be repeated.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 represents an exploded view of the sensor of the presentinvention.

FIG. 2 represents the sensor's base and those elements of the sensorwhich are applied directly to the base.

DESCRIPTION OF THE INVENTION

The construction of the electrochemical sensor with which the presentinvention is concerned is illustrated by FIG. 1. The sensor 34 is madeup of insulating base 36 upon which is printed in sequence (typically byscreen printing techniques) an electrical conductor pattern 38, anelectrode pattern (39 and 40) an insulating (dielectric) pattern 42 andfinally a reaction layer 44. The function of the reaction layer is toconvert glucose, or another analyte in the fluid test sample,stoichiometrically into a chemical species which is electrochemicallymeasurable, in terms of electrical current it produces, by thecomponents of the electrode pattern. The reaction layer typicallycontains an enzyme which reacts with the analyte to produce mobileelectrons on the electrode pattern and an electron acceptor such as aferricyanide salt to carry the mobile electrons to the surface of theworking electrode. The enzyme in the reaction layer can be combined witha hydrophilic polymer such as poly(ethylene oxide). The two parts 39 and40 of the electrode print provide the working 39 and counter 40electrodes necessary for the electrochemical determination of theanalyte which is the crux of the present invention. The working andcounter electrodes are configured in a manner such that the majorportion of the counter electrode is located downstream (in terms of thedirection of fluid flow along the flow path) from the exposed portion ofthe working electrode 39 a. This configuration offers the advantage ofallowing the test fluid to completely cover the exposed portion of theworking electrode for all cases in which an undetected partial fill hasoccurred. However, sub-element 40 a of the counter electrode ispositioned upstream from working electrode upper element 39 a so thatwhen an inadequate amount of fluid (such as blood) to completely coverthe working electrode enters the capillary space there will be formed anelectrical connection between counter electrode sub-element 40 a andexposed portion of the working electrode upper part 39 a due to theconductivity of the blood sample. However, the area of the counterelectrode which is available for contact by the blood sample is so smallthat only a very weak current can pass between the electrodes and hencethrough the current detector. By programming the current detector togive an error signal when any of the several error checking parametersis outside the tolerance range, the sensor device of the presentinvention actively advises the user that insufficient blood has enteredthe sensor's cavity and that another test should be conducted. The errorchecking parameters are derived from multiple current measurements. Onemain advantage of using error checking parameters, instead of directlychecking a weak current is that the short-fill detection works at bothhigh and low glucose levels. While the particular dimensions of theelectrodes are not critical, the area of the sub-element of the counterelectrode is typically less than about 10% than that of the workingelectrode and preferably less than about 6%. This element is made assmall as possible in view of the restraints of the screen printingprocess. It is also contemplated that reaction layer 44 can be removedfrom contact with sub-element 40 a of the counter electrode. This isaccomplished by producing a screen that does not print reagent ink overthe counter electrode sub-element 40 b and serves the purpose ofstarving the sub-element for reagent thereby not allowing it to functionas a proper counter electrode, so that an error condition is achieved inthe case of failure of the test fluid to contact the bulk of the counterelectrode 40. While sub-element 40 a is depicted as being physicallyconnected to, and therefore part of, the reference electrode 40, suchphysical connection is not critical. Such sub-element can be physicallydisconnected from the rest of the counter electrode provided that it isprovided with its own connector and the sensor is equipped with a thirdcontact to the detector.

The two parts 39 and 40 of the printed electrode provide the working andcounter electrodes necessary for the electrochemical determination ofanalyte. The electrode ink, which is about 14μ (0.00055″) thick,typically contains electrochemically active carbon. Components of theconductor ink are a mixture of carbon and silver which is chosen toprovide a low chemical resistance path between the electrodes and themeter with which they are in operative connection via contact with theconductive pattern at the fish-tail end of the sensor 45. The counterelectrode can be comprised of silver/silver chloride although carbon ispreferred. The function of the dielectric pattern is to insulate theelectrodes from the fluid test sample except in a defined area near thecenter of the electrode pattern to enhance the reproducibility of themeter reading. A defined area is important in this type ofelectrochemical determination because the measured current is dependentboth on the concentration of the analyte and the area of the reactionlayer which is exposed to the analyte containing test sample. A typicaldielectric layer 42 comprises a UV cured acrylate modified polymethanewhich is about 10μ (0.0004″) thick. The lid 46 which provides a concavespace 48, and which is typically formed by embossing a flat sheet ofdeformable material, is punctured to provide air vent 50 and joined tothe base 36 in a sealing operation. The lid and base can be sealedtogether by sonic welding in which the base and lid are first alignedand then pressed together between a vibratory heat sealing member orhorn and a stationary jaw. The horn is shaped such that contact is madeonly with the flat, non-embossed regions of the lid. Ultrasonic energyfrom a crystal or other transducer is used to excite vibrations in themetal horn. This mechanical energy is dissipated as heat in the plasticjoint allowing the bonding of the thermoplastic materials. The embossedlid and base can also be joined by use of an adhesive material on theunderside of the lid. The method of joining the lid and base are morefully described in U.S. Pat. No. 5,798,031 which is incorporated hereinby reference.

Suitable materials for the insulating base include polycarbonate,polyethylene terephthalate and dimensionally stable vinyl and acrylicpolymers as well as polymer blends such as polycarbonate/polyethyleneterephthalate and metal foil structures such as anylon/aluminum/polyvinyl chloride laminate. The lid is typicallyfabricated from a deformable polymeric sheet material such aspolycarbonate or an embossable grade of polyethylene terephthalate,glycol modified polyethylene terephthalate or a metal foil compositionsuch as an aluminum foil structure. The dielectric layer can befabricated from an acrylate modified polyurethane which is curable by UVlight or moisture or a vinyl polymer which is heat curable.

The construction of a sensor according to the present invention isaccomplished according to the following example:

EXAMPLE I

The base stock, typically of polycarbonate, is printed with various inksto form the electrodes 39 and 40 and then overcoated with a dielectriclayer 42 in a predetermined pattern designed to leave a desired surfaceof the electrode exposed to contact by the fluid test sample as itenters the space formed by the mating of lid 46 and base 36. Theparticular configuration of the dielectric layer 42 as depicted in FIG.1 in which opening 43 leaves the reagent layer in electricalcommunication with the electrodes 39 and 40 is designed to define theextent to which all of the conductive elements (working, reference andsub-element electrodes) are exposed to the test fluid. Along with theprinted conductive features, the dielectric layer defines the size ofeach of these elements. The electrodes are preferably printed so thatthe conductive and dielectric layers are close to 90 degrees to eachother. This helps in the tolerance stackup for building the sensorbecause it reduces the registration issues since as either printingshifts around the element, definition remains constant. The sensor baseof the present invention is also illustrated in FIG. 2 in which allelements on the base are shown in the same plane. The sensor's base 36has conductive element 38 on its surface which is in turn overcoatedwith working electrode 39 and counter electrode 40. Dielectric layer 42is not shown but instead the opening 43 in the dielectric layer is shownto illustrate the portions of working electrode 39 and counter electrode40 which are exposed. The sub-element of the counter electrode which isin electrical communication with the larger portion of the counterelectrode, designated as 40 b, functions in this embodiment to providean electrical conduction path with the working electrode such that thefluid can be detected as having reached the working electrode.Sufficient current will be provided to initiate the test sequence. Ifthe test fluid fails to fill the sensor cavity and contact the majorportion of the counter electrode, an error condition will be detectedand communicated to the user of the device.

A large number of sensors according to the present invention arefabricated from a rolled sheet of polycarbonate which has been unrolledto provide a flat surface. This sheet is referred to as the lid stocksince it serves as the source for a multiplicity of lids. There istypically placed a layer of thermoplastic adhesive on the underside ofthe lidstock after which concave areas 48 (FIG. 1) are embossed into thepolycarbonate sheet and various holes are punched into the sheet toprovide vent holes 50 and for registration and tracking before slitribbons of lidstock are rolled up. The base stock, typically ofpolycarbonate, is printed with various inks to form the electrodes andthen overcoated with the dielectric layer in a predetermined patterndesigned to leave a desired surface of the electrode exposed to thereaction layer 44 when it is printed over the dielectric layer.

The present invention introduces the advantage of providing anelectrochemical sensor in which the counter and working electrodes canbe configured so that in the event of a short fill, the result will beaffirmative as opposed to a neutral response, i.e. a failure of thedetector to give any signal. Thus, when the amount of test fluid whichenters the capillary space is sufficient to cover the sub-element of thecounter electrode 40 a, or 40 b in the preferred embodiment, and thatportion of the working electrode 39 a which lies upstream from the mainportion of the counter electrode 40, the detector will sense the valuesof error checking parameters derived from multiple current measurementsexceeding their tolerance limits if the working electrode is notcompletely covered with the test fluid. The detector can be connectedwith the reading means to provide an error signal which will alert theuser to the occurrence of a sort fill. The means of error checking areaccomplished by algorithmically programming the meter to detect theshort fill by measuring the current at a definite time period after thetest fluid has electrically connected the sub-element of the counterelectrode with the working electrode. The ratio of the currents for themeasurements is used to determine if the sensor has filled properly.Thus, a short fill is determined by employing the following steps:

-   -   a) making multiple current measurements at different time        periods when a driving potential is applied between the        electrodes;    -   b) converting the multiple current measurements into error        checking parameters; and    -   c) checking the values of the error checking parameters against        their corresponding tolerance limits to determine if a short        fill has occurred.

For example, in a sensor system which applied a 0.4 V potential for 10seconds after a blood sample is applied (known as the burn-off period),opens the circuit (OV potential) for 10 seconds (known as the waitperiod) and then applies a 0.4 V potential during the 10 second readperiod; the steps are carried out as follows:

Referring to Step A in the above paragraph, three current measurementsare made during the test sequence: 1) at the end of the burn-off perioddenoted as I_(r10); 2) at the 5 second during the read period denoted asI_(r5); and 3) at the end of the read period denoted as I_(r10).

Then in Step B, two parameters are determined from the three currentmeasurements. These two parameters are used to determine if the sensor'scapillary space has filled properly. The first parameter is the Decayfactor, which describes the shape of current time course. The secondparameter is the Read-to-Burn ratio that characterizes the magnitude ofinitial current in relation to the final current. The decay factor, k,is defined as:

$\begin{matrix}{k = \frac{{\ln\left( I_{r5} \right)} - {\ln\left( I_{r10} \right)}}{{\ln(10)} - {\ln(5)}}} & {{Eq}.\mspace{14mu} 1}\end{matrix}$Note: k characterizes how the current decays in a generalcurrent-glucose relationship I=c·G·t^(−k), where I is the current, c isa constant, G is the glucose concentration, and t is the time.The Read-to-Burn ratio, R/B is defined as:R/B=I _(r10) /I _(b10)  Eq. 2

In Step C, the values of these two parameters are checked against theirtolerance limits to determine if a short fill occurred. The tolerancelimits are not constant. They change as glucose level changes. Thetolerance-limit checking is described as Conditions 1 and 2 below. Thecriteria for a short fill are either Condition 1 or Condition 2 is true.

-   Condition 1 (Decay factor checking):    if |k−(a _(k1) +b _(k1) ·G)|>w _(k) is true when G≦d _(k1), or    if |k−(a _(k2) +b _(k2) ·G)|>w _(k) is true when d _(k1) <G≦d _(k2),    or    if |k−(a _(k3) +b _(k3) ·G)|>w _(k) is true when G>d _(k2)  Eq. 3    where a_(k1), a_(k2), a_(k3), b_(k1), b_(k2), b_(k3), w_(k), d_(k1),    d_(k2) and d_(k3) are predetermined constants, G is the glucose    measurement.-   Condition 2 (R/B ratio checking):    if |R/B−(a _(c1) +b _(c1) ·G)|>w _(c) is true when G≦d _(c), or    if |R/B−(a _(c2) +b _(c2) ·G)|>w _(c) is true when G>d _(c)  Eq. 4    where a_(c1), a_(c2), b_(c1), b_(c2), w_(c), and d_(c) are    predetermined constants, G is the glucose measurement.    The constants a_(k)'s, b_(k)'s, d_(k)'s and w_(k) in Eq. 3 are    predetermined experimentally:    -   Tests a large number of sensors at various glucose levels, G.    -   Calculates the decay factor, k, of each sensor from their I_(b5)        and I_(b10) currents.    -   Plots all the data points in a k vs. G chart.    -   Fits a 3-piece piecewise-linear line to the data points in the k        vs. G chart. These three pieces are a_(k1)+b_(k1)×G for        G≦d_(k1); a_(k2)+b_(k2)×G for G>d_(k1) and ≦d_(k2); and        a_(k3)+b_(k3)×G for G>d_(k2)    -   Add a tolerance width of ±w_(k) to the three lines so that the        band between the −w_(k) and +w_(k) is wide enough to enclose all        the normal data points in the chart.        The constants a_(c)'s, b_(c)'s, d_(c) and w_(c) in Eq. 4 are        also predetermined experimentally in the same way, on a R/B vs.        G chart.        A sample calculation is as follows:-   Step A—Make three current measurements of a sensor:    -   I_(b10)=505.1 nA, I_(r5)=656.5 nA, and I_(r10)=561.8 nA.-   Step B—Determine the value of the decay factor k and R/B ratio:    -   The decay factor and read-to-burn ratio were calculated from the        current measurements:    -   Decay factor

$k = {\frac{{\ln\left( I_{r5} \right)} - {\ln\left( I_{r10} \right)}}{{\ln(10)} - {\ln(5)}} = {\frac{{\ln(656.5)} - {\ln(561.8)}}{{\ln(10)} - {\ln(5)}} = 0.225}}$

-   -   Read-to-Burn ratio        R/B=I _(r10) /I _(b10)=561.8/505.1=1.11

-   Step C—Check against the tolerance limits:    The constants used in this example were:

-   a_(k1)=0.36, b_(k1)=−0.0002 dL/mg, w_(k)=0.13, and d_(k1)=100 mg/dL    The glucose reading from the sensor system is 22.9 mg/dL.    Condition 1 was true because of the first line in Eq. 3 was true.    if |k−(a _(k1) +b _(k1) ·G)|>w _(k) is true when G≦d _(k1)    |0.225−(0.36−0.0002·22.9)|=0.1304>0.13 is true when G=22.9≦d    _(k1)=100    No further check on Condition 2 was needed in this example, because    Condition 1 was already true.    Therefore, this sensor was determined as a short fill.

1. An electrochemical test sensor adapted to assist in determining theconcentration of an analyte in a fluid test sample, the sensorcomprising: a base that assists in forming an opening for introducingthe fluid test sample; a working electrode being coupled to the base; acounter electrode being coupled to the base, the counter electrode andthe working electrode being adapted to be in electrical communicationwith a detector of electrical current; and a sub-element being coupledto the base, a major portion of the counter electrode being locateddownstream relative to the opening and at least a portion of the workingelectrode, the sub-element being located upstream relative to theworking electrode such that when electrical communication occurs betweenonly the sub-element and the working electrode there is insufficientflow of electrical current through the detector to determine theconcentration of the analyte in the fluid test sample.
 2. The sensor ofclaim 1 further comprising a cover adapted to be coupled to the base toform a capillary space, the capillary space having an opening forintroducing the fluid test sample therein, the capillary space forming aflow path for the fluid test sample, the working electrode and thecounter electrode being situated in the flow path.
 3. The sensor ofclaim 2, wherein the cover includes an air vent.
 4. The sensor of claim1 further comprising a reaction layer located on the surface of at leastthe working electrode, the reaction layer comprising an enzyme adaptedto react with the analyte to produce electrons, the electrons beingadapted to be transferred to the working electrode.
 5. The sensor ofclaim 1, wherein the area of the sub-element is less than 10% of thearea of the working electrode.
 6. The sensor of claim 1, wherein thesub-element and the counter electrode are physically connected.
 7. Amethod of determining whether a sufficient quantity of a fluid testsample has been introduced to an electrochemical test sensor, the methodcomprising the acts of: providing the electrochemical test sensoradapted to assist in determining the concentration of an analyte in afluid test sample, the sensor comprising a base that assists in formingan opening for introducing the fluid test sample, a working electrodebeing coupled to the base, a counter electrode being coupled to thebase, and a sub-element being coupled to the base, a major portion ofthe counter electrode being located downstream relative to the openingand at least a portion of the working electrode, the sub-element beinglocated upstream relative to the working electrode; introducing thefluid test sample to the test sensor; and determining whether asufficient quantity of the fluid test sample has been introduced to theelectrochemical test sensor and, if not, notifying a user that aninsufficient quantity of the fluid test sample has been introduced. 8.The method of claim 7 further comprising, prior to the act ofdetermining whether a sufficient quantity of the fluid test sample hasbeen introduced to the electrochemical test sensor, measuring current ata plurality of time periods to obtain a plurality of currentmeasurements.
 9. The method of claim 8 further comprising providing acurrent to initiate a test sequence prior to the act of measuringcurrent at a plurality of time periods.
 10. The method of claim 8further comprising, prior to the act of measuring current at a pluralityof time periods, the acts of: converting the plurality of currentmeasurements into a plurality of error checking parameters; andcomparing the plurality of error checking parameters with theircorresponding tolerance ranges.
 11. The method of claim 10 wherein theact of notifying a user that an insufficient quantity of the fluid testsample has been introduced occurs when one or more of the plurality oferror checking parameters are outside of their corresponding toleranceranges.
 12. The method of claim 11, wherein one or more of the pluralityof error checking parameters is outside of its corresponding tolerancerange when the fluid test sample fails to contact a major portion of thecounter electrode.
 13. The method of claim 10, wherein the plurality oferror checking parameters is derived from the plurality of currentmeasurements.
 14. The method of claim 7, wherein the act of applying thefluid test sample to the sensor electrically connects the sub-element tothe working electrode.
 15. The method of claim 7, wherein the act ofnotifying a user that an insufficient quantity of the fluid test samplehas been introduced includes providing an error signal.
 16. The methodof claim 7, wherein the test sensor further comprises a cover adapted tobe coupled to the base to form a capillary space, the capillary spacehaving an opening for introducing the fluid test sample therein, thecapillary space forming a flow path for the fluid test sample, theworking electrode and the counter electrode being situated in the flowpath.
 17. The method of claim 16, wherein the cover includes an airvent.
 18. The method of claim 7, wherein the test sensor furthercomprises a reaction layer located on the surface of at least theworking electrode, the reaction layer comprising an enzyme adapted toreact with the analyte to produce electrons, the electrons being adaptedto be transferred to the working electrode.
 19. The method of claim 7,wherein the area of the sub-element is less than 10% of the area of theworking electrode.
 20. The method of claim 7, wherein the sub-elementand the counter electrode are physically connected.